Transcriptomic analyses of joint tissues during osteoarthritis development in a rat model reveal dysregulated mechanotransduction and extracellular matrix pathways.

OBJECTIVE: Transcriptomic changes in joint tissues during the development of osteoarthritis (OA) are of interest for the discovery of biomarkers and mechanisms of disease. The objective of this study was to use the rat medial meniscus transection (MMT) model to discover stage and tissue-specific transcriptomic changes. DESIGN: Sham or MMT surgeries were performed in mature rats. Cartilage, menisci and synovium were scored for histopathological changes at 2, 4 and 6 weeks post-surgery and processed for RNA-sequencing. Differentially expressed genes (DEG) were used to identify pathways and mechanisms. Published transcriptomic datasets from animal models and human OA were used to confirm and extend present findings. RESULTS: The total number of DEGs was already high at 2 weeks (723 in meniscus), followed by cartilage (259) and synovium (42) and declined to varying degrees in meniscus and synovium but increased in cartilage at 6 weeks. The most upregulated genes included tenascins. The 'response to mechanical stimulus' and extracellular matrix-related pathways were enriched in both cartilage and meniscus. Pathways that were enriched in synovium at 4 weeks indicate processes related to synovial hyperplasia and fibrosis. Synovium also showed upregulation of IL-11 and several MMPs. The mechanical stimulus pathway included upregulation of the mechanoreceptors PIEZO1, PIEZO2 and TRPV4 and nerve growth factor. Analysis of data from prior RNA-sequencing studies of animal models and human OA support these findings. CONCLUSION: These results indicate several shared pathways that are affected during OA in cartilage and meniscus and support the role of mechanotransduction and other pathways in OA pathogenesis.

Dynamic shear analysis: a novel method to determine mechanical integrity of normal and torn human acetabular labra: Implications for prediction of outcome of repair.

OBJECTIVES: The aim of this study was to investigate the structural integrity of torn and non-torn human acetabular labral tissue. METHODS: A total of 47 human labral specimens were obtained from a biobank. These included 22 torn specimens and 25 control specimens from patients undergoing total hip arthroplasty with macroscopically normal labra. The specimens underwent dynamic shear analysis using a rheometer to measure storage modulus, as an indicator of structural integrity. RESULTS: There was a significant difference in the storage modulus between torn (mean modulus = 2144.08 Pa) and non-torn (3178.1 Pa) labra (p = 0.0001). CONCLUSION: The acetabular labrum of young patients with a tear has significantly reduced structural integrity compared with a non-torn labrum in older patients with end-stage osteoarthritis. This study contributes to the understanding of the biomechanics of labral tears, and the observation of reduced structural integrity in torn labra may explain why some repairs fail. Our data demonstrate that labral tears probably have a relatively narrow phenotype, presenting a basis for further investigations that will provide quantifiable data to support their classification and a means to develop a standardized surgical technique for their repair. This study also demonstrates the value of novel biomechanical testing methods in investigating pathological tissues of orthopaedic interest.Cite this article: A. K. Woods, J. Broomfield, P. Monk, F. Vollrath, S. Glyn-Jones. Dynamic shear analysis: a novel method to determine mechanical integrity of normal and torn human acetabular labra: Implications for prediction of outcome of repair. Bone Joint Res 2018;7:440-446. DOI: 10.1302/2046-3758.77.BJR-2017-0282.R2.

A Novel Piperazine-Based Drug Lead for Cryptosporidiosis from the Medicines for Malaria Venture Open-Access Malaria Box.

Cryptosporidiosis causes life-threatening diarrhea in children under the age of 5 years and prolonged diarrhea in immunodeficient people, especially AIDS patients. The standard of care, nitazoxanide, is modestly effective in children and ineffective in immunocompromised individuals. In addition to the need for new drugs, better knowledge of drug properties that drive in vivo efficacy is needed to facilitate drug development. We report the identification of a piperazine-based lead compound for Cryptosporidium drug development, MMV665917, and a new pharmacodynamic method used for its characterization. The identification of MMV665917 from the Medicines for Malaria Venture Malaria Box was followed by dose-response studies, in vitro toxicity studies, and structure-activity relationship studies using commercial analogues. The potency of this compound against Cryptosporidium parvum Iowa and field isolates was comparable to that against Cryptosporidium hominis Furthermore, unlike nitazoxanide, clofazimine, and paromomycin, MMV665917 appeared to be curative in a NOD SCID gamma mouse model of chronic cryptosporidiosis. MMV665917 was also efficacious in a gamma interferon knockout mouse model of acute cryptosporidiosis. To determine if efficacy in this mouse model of chronic infection might relate to whether compounds are parasiticidal or parasitistatic for C. parvum, we developed a novel in vitro parasite persistence assay. This assay suggested that MMV665917 was parasiticidal, unlike nitazoxanide, clofazimine, and paromomycin. The assay also enabled determination of the concentration of the compound required to maximize the rate of parasite elimination. This time-kill assay can be used to prioritize early-stage Cryptosporidium drug leads and may aid in planning in vivo efficacy experiments. Collectively, these results identify MMV665917 as a promising lead and establish a new method for characterizing potential anticryptosporidial agents.

Histological and mechanical differences in the skin of patients with rectal prolapse.

INTRODUCTION: It is still an enigma that some patients develop rectal prolapse whilst others with similar risk factors do not. Biomechanical assessment of the skin may provide further insight into the aetiology of this complex condition. Elastin fibres are an abundant and integral part of many extracellular matrices and are especially critical for providing the property of elastic recoil to tissues. The significance of elastin fibres is clearly reflected by the numerous human conditions in which a skin phenotype occurs as a result of elastin fibre abnormalities. METHOD: Between January and June 2013, skin specimens were obtained prospectively during surgery on 20 patients with rectal prolapse and 21 patients without prolapse undergoing surgery for other indications. Expression levels of elastin in the skin were measured by Orcein staining, and Image J. Tensile tests were performed using the Zwick Roell device, with custom ceramic clamps. For statistical analysis, Student's t test was used. RESULTS: Histological analysis of prolapse vs control showed percentage dermal elastin fibres of 9 vs 5.8 % (p = 0.001) in males and 6.5 vs 5.3 % (p = 0.05) in females. Patients with more severe prolapse (external) had a significantly (p = 0.05) higher percentage dermal elastin fibres 6.9 vs 6.1 % than internal prolapse. Young's modulus of patients with prolapse was lower in males (3.3 vs 2.8, p = 0.05) and females (3.1 vs 2.7, p = 0.05). CONCLUSION: Patients with prolapse have a higher concentration of elastin fibres in the skin, and these differences are quantitatively demonstrated through mechanical testing. This suggests that the aetiology may be a result of a dysfunction of elastin fibre assembly.

Distal-less 3 haploinsufficiency results in elevated placental oxidative stress and altered fetal growth kinetics in the mouse.

Distal-less 3 (Dlx3)(-/-) mice die at E9.5 presumably due to an abnormal placental phenotype including reduced placental vasculature and secretion of placental growth factor. To examine the role of Dlx3 specifically within the epiblast, Dlx3 conditional knockout mice were generated using an epiblast-specific Meox2(CreSor) allele. Dlx3(-/fl), Meox2(CreSor) animals were born at expected frequencies and survived to weaning providing indirect evidence that loss of Dlx3 within the trophoectoderm plays a critical role in fetal survival in the Dlx3(-/-) mouse. We next examined the hypothesis that loss of a single Dlx3 allele would have a negative impact on placental and fetal fitness. Dlx3(+/-) mice displayed reduced fetal growth beginning at E12.5 compared with Dlx3(+/+) controls. Altered fetal growth trajectory occurred coincident with elevated oxidative stress and apoptosis within Dlx3(+/-) placentas. Oral supplementation with the superoxide dismutase mimetic, Tempol, rescued the fetal growth and placental cell death phenotypes in Dlx3(+/-) mice. To determine the potential mechanisms associated with elevated oxidative stress on the Dlx3(+/-) placentas, we next examined vascular characteristics within the feto-placental unit. Studies revealed reduced maternal spiral artery luminal area in the Dlx3(+/-) mice receiving water; Dlx3(+/-) mice receiving Tempol displayed maternal spiral artery luminal area similar to control Dlx3(+/+) mice. We conclude that reduced Dlx3 gene dose results in diminished fetal fitness associated with elevated placental cell oxidative stress and apoptosis coincident with altered vascular remodeling. Administration of antioxidant therapy ameliorated this feto-placental phenotype, suggesting that Dlx3 may be required for adaptation to oxidative stresses within the intrauterine environment.

Maximal enzyme activities, and myoglobin and glutathione concentrations in heart, liver and skeletal muscle of the Northern Short-tailed shrew (Blarina brevicauda; Insectivora: Soricidae).

We measured the enzymes of glycolysis, Krebs Cycle, beta-oxidation and electron transport in the heart, liver and skeletal muscle of the Northern Short-tailed Shrew, Blarina brevicauda. Additionally, we measured the amount of myoglobin in skeletal and heart muscle as well as the concentration of glutathione in heart. The picture that emerges is of an aerobically well-endowed animal with constrained anaerobic capacity as indicated by small activities of glycolytic enzymes and creatine kinase. Lipid metabolism and amino acid transamination, as well as gluconeogenesis, are predominant in processing carbon resources and probably reflect the large contribution lipid and protein make to the diet of this carnivore. The citrate synthase activity is the largest of any reported value for vertebrate heart (250 U/g). The additional, very active cytochrome c oxidase activity (220 U/g) and large myoglobin concentrations (8 mg/g) in heart are clearly the underpinnings of the rapid metabolic rates reported for small insectivores. The potential for generation of reactive oxygen species must be great since the total glutathione concentration (165 mumol/g) is 300-fold greater in shrew hearts than in hearts of rats.